Rat pheochromochytoma tyrosine hydroxylase was cloned, expressed in E.coli, and subsequently purified to homogeneity. The pure recombinant hydroxylase exhibited the same kinetic behavior as that of the activated native enzyme. This activation was reversed, however, when the enzyme was treated with one of the catecholamine products, dopamine. Dopamine was shown to bind with high-affinity to the recombinant enzyme and to cause a significant change i the kinetic as well as spectral properties of the enzyme. The results explain the activation of the recombinant enzyme by the absence of dopamine in E.coli, and the consequent lack of dopamine-mediated down regulation of the hydroxylase. Furthermore, the date identify dopamine binding and inhibition as an essential eukaryotic post-translational modification that serves to regulate tyrosine hydroxylase activity. Several deletion mutants of tyrosine hydroxylase were expressed in E.coli and partially purified for further characterization. The mutant enzyme forms displayed different levels of catalytic activity and very distinct kinetic properties, which were correlated to the size and location of the deletion. The results have helped to redefine the boundaries of the catalytic domain and to clarify th role of the N-terminus in directing substrate specificy, cofactor binding and product inhibition. Other studies have continued to explore the mechanism of tyrosine hydroxylase phosphorylation and dephosphorylation in intact rat striatal synaptosomes. Earlier evidence unveiled a pathway of dephosphorylation for tyrosine hydroxylase which was markedly stimulated by BH4 in situ. Current studies are attempting to identy the specific phosphoamino acid target sites for BH4.